The soil microbe Bacillus thuringiensis (B.t.) is a Gram-positive, spore-forming bacterium characterized by parasporal crystalline protein inclusions. These inclusions often appear microscopically as distinctively shaped crystals. The B.t. microbes produce a variety of toms. A mouse-lethal ct-exotoxin is produced by certain isolates of this species, as well as two types of hemolysins, a nucleotide .beta.-exotoxin and .delta.-endotoxins. Delta-endotoxins are derived from the parasporal crystalline protein inclusions and are highly toxic to pests and specific in their toxic activity. The B.t. toxins have been commercially exploited for their use in pest control. Certain B.t. endotoxin genes have been isolated and sequenced, and recombinant DNA-based B.t. products have been produced and approved for use. In addition, with the use of genetic engineering techniques, new approaches for delivering B.t. endotoxins to agricultural environments are under development, including the use of plants genetically engineered with endotoxin genes for insect resistance and the use of stabilized intact microbial cells as B.t. endotoxin delivery vehicles (Gaertner, F.H., L. Kim [1988]TIBTECH 6:S4-S7).
Until the last 10 years, commercial use of B.t. pesticides has been largely restricted to a narrow range of lepidopteran (caterpillar) pests. Preparations of the spores and crystals of B. thudngiensis subsp. kurstaki have been used for many years as commercial insecticides for lepidopteran pests. For example, B. thuringiensis var. kurstaki HD-1 produces a delta-endotoxin which is toxic to the larvae of a number of lepidopteran insects.
In recent years, however, investigators have discovered B.t. pesticides with specificities for a much broader range of pests. For example, other species of B.t., namely israelensis and san diego (a.k.a.B.t. tenebdonis, a.k.a. M-7), have been used commercially to control insects of the orders Diptera and Coleoptera, respectively (Gaertner, F.H. [1989]"Cellular Delivery Systems for Insecticidal Proteins: Living and Non-Living Microorganisms," in Controlled Delivery of Crop Protection Agents, R.M. Wilkins, ed., Taylor and Francis, New York and London, 1990, pp. 245-255). See also Couch, T.L. (1980) "Mosquito Pathogenicity of Bacillus thudngiensis var. israelensis," Developments in Industrial Microbiology 22:61-76; Beegle, C.C., (1978) "Use of Entomogenous Bacteria in Agroecosystems," Developments in Industrial Microbiology 20:97-104. Krieg, A., A.M. Huger, G.A. Langenbruch, W. Sctmetter (1983) Z. ang. Ent. 96:500-508, describe a B.t. isolate named Bacillus thuringiensis var. tenebrionis, which is reportedly active against two beetles in the order Coleoptera. These are the Colorado potato beetle, Leptinotarsa decemlineata, and Agelastica alni.
There have been published reports concerning the use of Bacillus thudngiensis preparations for the control of acarid pests or mites. These publications are as follow:
Royalty, R.N., F.R. HaH, R.A.J. Taylor (1990) "Effects of thuringiensin on Tetranychus urticae (Acari: Tetranychidae) mortality, fecundity, and feeding," J. Econ. Entomot 83:792-798. PA0 Neal, J.W., R.K. Lindquist, K.M. Gott, M.L. Casey (1987) "Activity of the themostable beta-exotoxin of Bacillus thudngiensis Berliner on Tetranychus urticae and Tetranychus cinnabarinus,"J. Agric. Entomol. 4:33-40. PA0 Vlayen, P., G. Impe, R. Van Semaille (1978) "Effect of a commercial preparation of Bacillus thuringiensis on the spider mite Tetranychus urticae Koch. (Acari: Tetranychidae)," Mededelingen 43:471-479.
In the above published studies, the active ingredient in the B.t. preparations was .beta.-exotoxin (also called thuringiensin).
The major focus for commercial use of B.t. toxins is on the .delta.-endotoxins from the parasporal crystalline protein inclusions. Recently, new subspecies of B.t. have been identified, and genes responsible for active .delta.-endotoxin proteins have been isolated (Hofte, H., H.R. Whiteley [1989]Microbiological Reviews 52(2):242-255). Hofte and Whiteley classified B.t. crystal protein genes into 4 major classes. The classes were CryI (Lepidoptera-specific), CrylI (Lepidoptera- and Diptera-specific), CrylII (Coleoptera-specific), and CryIV (Diptera-specific). The discovery of strains specifically toxic to other pests has been reported. (Feitelson, J.S., J. Payne, L. Kim [1992]Bio/Technology 10:271-275).
The cloning and expression of a B.t. crystal protein gene in Escherichia coli has been described in the published literature (Schnepf, H.E., H.R. Whitely [1981]Proc. Natl. Acad. Sci. USA 78:2893-2897). U.S. Pat. No. 4,448,885 and U.S. Pat. No. 4,467,036 both disclose the expression of B.t. crystal proteins in E. coli. U.S. Pat. Nos. 4,797,276 and 4,853,331 disclose B.thuringiensis strain san diego (a.k.a.B.t. tenebrionis, a.k.a. M-7) which can be used to control coleopteran pests in various environments. U.S. Pat. No. 4,849,217 discloses isolates of B.t. which have activity against a coleopteran pests, the alfalfa weevil, while U.S. Pat. No. 4,948,734 discloses certain isolates of B.t. which have activity against nematodes. U.S. Pat. No. 5,093,120 discloses the use of these B.t. isolates to control nematode infections in animals and plants. Many other patents have issued for new B.t. isolates and new uses of B.t. isolates. The discovery of new B.t. isolates and new uses of known B.t. isolates remains an empirical, unpredictable art.